Lactic acid extraction and purification process

ABSTRACT

The subject of the invention is a process for extracting pure lactic acid from fermentation liquors by ion exchange chromatography on a strongly acidic cation exchanger, preferably in H +   form. In a first step, the NH 4  lactate coming from the fermentation is converted into the free acid by genuine ion exchange. This conversion is preferably effected on a weakly acidic cation exchanger in H +   form.

BACKGROUND OF THE INVENTION

The invention is directed to a process for the separation andpurification of lactic acid from salt-containing andcarbohydrate-containing substrates (fermentation solution) from whichcoarsely dispersed and lipophilic impurities have been removed.

Industrial production of lactic acids, especially where pure L(+)-lacticacid and D(-)-lactic acid are to be extracted, is presently carried outpredominantly by biotechnological processes. The production process canbe divided into:

1) the actual production of lactic acid by fermentation of acarbohydrate-containing medium;

2) the working up (downstream processing) of the fermentation solutionto form pure acid.

Industry has adopted a large number of strains of microorganisms for theproduction of lactic acid. The most important of these are thehomofermentative lactic acid bacteria of the genera lactobacillus,streptococcus and pediococcus. However, these genera reach their maximumproductivity only within a very narrow pH range.

Therefore, it is necessary during fermentation not only to maintain aconstant optimum temperature for the selected organism, but also tomaintain the required pH at a constant value. For this reason,neutralizing agents such as alkali hydroxide, calcium carbonate, milk oflime or ammonia water are added to the mash before and/or duringfermentation so as to prevent over-acidification and to maintain aconstant pH of 5.5 to 6.5.

Thus, the main component contained in the fermentation mash is the saltof lactic acid (e.g., NH₄ lactate or Ca lactate) in addition to a littlefree acid, as well as unconverted starter materials (e.g., sugar), heavymetals, coloring matter, metabolic by-products (e.g., acetic or ethanoicacid), cells and cell fragments of the microorganisms, and inorganicsalts.

Therefore, direct use of the solution coming from fermentation is notpossible and further processing steps are required to extract the pure,free lactic acid.

A number of methods are described for the separation and extraction offree lactic acid from the fermentation mash.

The simplest commercial method of purification, and that most oftenapplied by manufacturers, is precipitation of the lactic acid as calciumlactate. In this method, at the end of the fermentation process the mashis first heated to approximately 80°-90° C. and the pH is increased to10-11. In this step, the microorganisms are destroyed, the proteins arecoagulated and the formed calcium lactate is dissolved. After allinsoluble components have been separated, the mash is acidified withsulfuric acid to liberate the lactic acid from its salt. In order toremove the iron and copper ions introduced especially as a result ofcorrosion, as well, sodium hexacyanoferrate (II) or calciumhexacyanoferrate (II) is added and the precipitated ferrocyanide salts,together with the calcium sulfate, are separated by means of a rotaryfilter or filter press. Coloring components are removed by activatedcharcoal. The obtained dilute acid is then concentrated to lactic acidof approximately 80% strength, small amounts of resulting volatile acidsbeing removed at the same time.

In improved precipitation processes, the crude lactic acid is firstdecolorized with activated charcoal and subsequent purification stepscan be carried out by means of cation exchangers for complete removal ofany remaining salts. The cation-free solution can then be evaporated orcrystallized immediately or can be guided through an anion exchanger inorder to remove any remaining foreign anions, mainly sulfate ions andchloride ions (DD-PS 6740).

For a further improvement in quality, especially with respect to odorand flavor, an oxidative treatment with hydrogen peroxide or potassiumpermanganate is frequently also carried out subsequently (CARLOSBELLAPART VILA, 1964, ES 297969).

However, all precipitation methods have grave disadvantages. Apart fromthe relatively high technical costs, the primary disadvantage consistsin a material loss of up to 20% of the lactic acid due to theprecipitation and crystallization processes (HEDING, L. G., Biotechm.Bioeng. 17, 1975, 1363-1364). Moreover, these methods require largequantities of auxiliary chemicals. Since the lactic acid occurs in mostcases as calcium lactate which must first be converted to free acid withsulfuric acid and an equivalent amount of gypsum or calcium sulfate, thecosts for disposing of large amounts of calcium sulfate are added to thecost for lime and sulfuric acid.

The grade of lactic acid which can be produced by this "precipitationmethod" is edible-grade lactic acid and is accordingly only suitable forthe foodstuffs industry. However, the pharmaceutical industry requireslactic acid of higher purity. Even higher criteria for purity arerequired for plastics produced from lactic acid by polymerization. Inparticular, this requires a total absence of carbohydrates. Therefore,there has been no lack of attempts to find other methods for extractingthe lactic acid from the fermentation solution in order to producehigher grades of lactic acid (pharmaceutical-grade lactic acid,plastic-grade lactic acid).

One possibility for producing pharmaceutical-grade lactic acid is steamdistillation with superheated steam under vacuum. Lactic acid, as iswell-known, has a very low volatility with steam at 100° C. However, thesteam volatility can be considerably increased by using superheatedsteam in a temperature range of 160°-200° C. Based on these results,methods for purification of lactic acid by steam distillation have beenworked out, e.g., as described in patents DK 83589 (1957) or CS 97136(1960). However, this method--by far the oldest--for production ofpharmaceutical-grade lactic acid has not been successful in practicesince this purification process is much too costly due to the relative"nonvolatility" of the lactic acid.

Liquid-liquid extraction of lactic acid with organic solvents, however,has been more successful. In principle, the procedure in this methodconsists in that the fermentation solution which has been freed ofbiological matter is acidified with sulfuric acid, the precipitatedcalcium sulfate is removed by filtration and, finally, decolorizing iscarried out with activated charcoal and salts are removed by ionexchange. The crude lactic acid solution produced in this way is thenconcentrated under vacuum to a determined concentration and is broughtinto contact with an organic solvent in a countercurrent extractioncolumn. The lactic acid can then be extracted from the organic phaseeither by backextraction with water or by distilling off the organicsolvent. Further treatment of the pure lactic acid solution withactivated charcoal and ion exchangers is often required after theextraction process before it can be concentrated to the conventionalcommercial concentration of 80%.

A process of this kind is described, e.g., by JENEMANN (1933) in U.S.Pat. No. 1,906,068, where isopropyl ether is used as a solvent.

An extraction process using nitroparaffin as the organic phase isproposed in TINDALL (1940), U.S. Pat. No. 2,223,797.

Also, in more recent times there has been no shortage of attempts toimprove the process for obtaining lactic acid by extraction.

For example, DE-OS 3415141 proposes an extraction process in whichbutanols or pentanols are used as solvents. The characteristic featurein this method consists in that the liquor containing calcium lactate isacidified with sulfuric acid immediately after fermentation and theobtained suspension which contains calcium sulfate and biomass as solidsis brought into contact with the solvent directly in a pulsedcountercurrent column outfitted with built-in hydrophobic pieces (e.g.,made of Teflon). After the extraction of the aqueous suspension by thesolvent, which is preferably carried out at a temperature of 70° C., asolids-containing aqueous phase and a solids-free organic phase areremoved from the column. The lactic acid dissolved in the organic phaseis finally converted completely into the lactic acid ester by distillingthe reaction water at 60°-140° C. (possibly under vacuum). This lacticacid ester can be obtained in pure form by vacuum distillation and is avaluable intermediate product. The esters can be split again into lacticacid and alcohol, as is well-known, so that highly purepharmaceutical-grade lactic acid can be obtained.

A great disadvantage in all extraction methods consists in that most ofthe organic solvents used for lactic acid have only a very lowdistribution coefficient so that very large quantities of organicsolvents are required.

However, the distribution coefficient for lactic acid can besubstantially improved when a mixture of organic solvents with atertiary amine is used for extraction. A purification process based onthis principle is described, for example, in U.S. Pat. No. 4,698,303(1987) in which a mixture of approximately 60%-75% isobutyl heptylketone and 25%-40% Adogen 364 was proven especially effective as anextraction medium. Adogen 364 is the trade name (Sherex Co.) of amixture of long-chain (C8-C10) tertiary amines.

However, certain difficulties arise in this purification process inrecovering the lactic acid from the organic phase, since this can onlybe carried out by backextraction with a basic solution (preferablyammonium hydroxide). This means that additional purification steps arerequired after extraction.

Although the lactic acid obtained by liquid-liquid extraction issubstantially free of ash, it does contain other impurities stemmingfrom the raw material. Very pure starter mashes as well as additionaltreatments, e.g., with activated charcoal, oxidizing agents and ionexchangers, are required to obtain pharmaceutical-grade lactic acid bythis method (VICKROY, T. B., Lactic Acid in. ComprehensiveBiotechnology, Vol. 3, 761-776, Pergamon Press) (PECKHAM, G. T., 1944,Chem. Eng. News, 22, 440-443).

Therefore, the method most frequently applied for the production ofpharmacopeia lactic acid is esterification of the lactic acid with lowalcohols (usually methanol) and subsequent separation of the esters byfractional distillation. Numerous methods for the purification of lacticacid by esterification are described in the literature. In a part ofthis process the crude lactic acid which is reduced to a determinedconcentration, and to which is generally added an acid catalyst, isexposed to the action of alcohol vapors. For the most part, lactic acidis separated from the escaping vapor mixture as esters of theaccompanying substances. The surplus alcohol is separated and fed backin a subsequent rectifying column. The methyl lactate can finally behydrolyzed again with water to form methanol and lactic acid.

The esterifying reaction of concentrated lactic acid with methanol inthe presence of an acid catalyst does not present any difficulties withrespect to purification. Thus, DE-OS 1912730 describes a process for theproduction of lactic acid methyl ester in the presence of an acidic ionexchanger, wherein the ester is obtained in a yield of 82 percent byfractional vacuum distillation.

Purification of the esters becomes more difficult when starting from adiluted aqueous lactic acid solution because the ester can be hydrolyzedagain very easily in the presence of water. In U.S. Pat No. 2,350,370,although diluted aqueous lactic acid is esterified with an acidcatalyst, the distilled ester is saponified again immediately in orderto purify the lactic acid.

DE-OS 3214697 proposes a process for continuous purification of lacticacid methyl esters in which the ester, which is produced byesterification of a diluted lactic acid with an acid catalyst, is firstconcentrated by partial condensation of the gas mixture occurring duringesterification and by subsequent vacuum distillation, and the crudelactic acid methyl ester remaining in the sump or bottom of the firstseparation column which contains only essentially small amounts oflactic acid is guided into a second separation column for completepurification.

Even though the "esterification method" is currently the only usablemethod for producing pharmacopeia-grade lactic acid, it still has thedisadvantage that large amounts of organic solvent must also be used,which poses a considerable safety hazard and risk to the environment.

For this reason there was an intensification of the search foralternative methods not having the disadvantages mentioned above.

Thus, a number of patents are known which propose that the organic acidsbe separated by electrodialysis. AT 290441, for example, proposes aprocess for purification of lactic acid in which a lactic acid which isfree from "unpleasant odor and taste" is obtained by a combination ofelectrodialysis and extraction. In this process, a crude lactic acidsolution which has been concentrated to approximately 20% is subjectedto electrodialysis treatment and the dialyzed solution is then extractedwith an organic solvent (isopropyl ether is recommended). The lacticacid is obtained from the organic phase by backextraction with water.

A process for purifying lactic acid is suggested in EP 0393818, in whichthe lactic acid salt (e.g., NH₄ lactate) contained in the fermentationliquor is first separated by conventional electrodialysis. The lacticacid salt extracted in this way is then directed to a secondelectrodialyzer which is outfitted with bipolar membranes and in whichthe lactic acid salt is separated into the free acid and itscorresponding base by hydrolysis. Finally, the lactic acid solution isguided through a strongly basic and a strongly acidic ion exchange resinin order to remove any anions and cations which may be present. A lacticacid of high purity is obtained in this way.

However, processes using electrodialysis have the disadvantage that theyrequire large amounts of electrical energy on the one hand, whichrenders the process very expensive, and additional purification steps,e.g., ion exchange, on the other hand in order to produce highly purelactic acid.

Another alternative method for extracting and/or purifying carboxylicacids produced by fermentation is the ion exchange method with acidicand/or basic ion exchange resins.

DD-PS 203533 describes an ion exchange process for extracting carboxylicacids and hydroxycarboxylic acids from their solutions containingforeign salts, in which the salts are first transformed into acids viastrongly acidic cation exchangers in H⁺ form. In so doing, a mixture ofcarboxylic acids and foreign acids is obtained. A weakly basic anionexchanger which is first present in the form of a base or hydroxyl isloaded with a small concentrated fraction of this mixture, i.e.,substantially converted to the form of carboxylic acid. Next, a morehighly concentrated fraction of the acid mixture obtained in thedecationization step is guided via the acid-charged ion exchanger. Theacid to be extracted passes freely through the resin bed and only thestronger foreign anions (chloride is mentioned) bind to the resin by ionexchange.

EP 0135728 describes a process for "isolating enzymatically producedcarboxylic acids" in which the solution which is advantageously producedby continuous fermentation runs through a "desorber" filled with a"polymer with tertiary amino groups" which selectively adsorbscarboxylic acids. The liquid exiting from the "desorber" which isextensively free of carboxylic acids is guided back into the reactoragain. After exhausting a "desorber", the reaction solution is guided tothe next desorber and the carboxylic acid is eluted from the exhausteddesorber by means of a polar solvent, e.g., methanol. The separation ofthe carboxylic acid from the eluate is carried out by known methods,e.g., by distillation in the case of volatile eluting agents.

Other processes for separating lactic acid by ion exchange are suggestedin U.S. Pat. No. 3,202,705 and JP 91183487. In these processes, afterseparation of the precipitated CaSO₄, the fermentation liquor which isacidified with sulfuric acid is first guided through a strongly acidiccation exchanger in H⁺ form and then through a basic anion exchanger. Inthis way, a "colorstable" lactic acid is obtained, at least in the U.S.patent.

The great disadvantage in all methods in which a "genuine" ion exchangetakes place is the required cost for regenerating the resins.

Therefore, chromatography processes on basic anion exchangers aredescribed as the most up-to-date alternative method for extracting andpurifying organic acids.

For example, EP 0324210 proposes a purification process in which acitric acid produced by fermentation is purified by adsorption atneutral, nonionic, macroreticular, water-insoluble resins or at weaklyor strongly basic anion exchangers. Water, a mixture of water andacetone, or a diluted sulfuric acid solution are used as eluent. Thismethod is capable of separating salts and carbohydrates from the citricacid.

The process proposed in EP-OS 0377430 works on precisely the sameprinciple. In this method, basic ion exchangers are also suggested forchromatographic isolation and/or purification of acids. The chiefdifference compared to EP 0 324 210 consists in that this method can beused to separate and/or purify not only citric acid, but also otherinorganic acids (e.g., phosphoric acid) and organic acids (tartaricacid, malic acid or lactic acid).

However, all ion exchange chromatography or IEC methods using basicanion exchangers or nonionic adsorber resins have the great disadvantagethat the acids retained on the resin by adsorption show a high degree of"tailing" when eluted with water or with diluted sulfuric acid, whichresults in an intensive dilution of the eluted acid. Further, it isimpossible with this method to separate acids having similar pKs values,e.g., lactic acid and ethanoic acid.

SUMMARY OF THE INVENTION

The object of the present invention is to extract lactic acid from salt-and carbohydrate-containing substrates (fermentation mashes) in a simpleand reliable manner.

This object is met, according to the invention, in that the separationand purification process is carried out in two steps:

a) in the first step, the salts which may be present in the fermentationsolution, principally the salt of lactic acid, are converted into freeacids by means of genuine ion exchange in one or more "preliminarycolumns", and

b) in the second step, the free lactic acid is separated from the restof the acids, carbohydrates and other impurities present in thefermentation solution by chromatography at strongly acidic ionexchangers in one or more "separation columns".

This process provides the following advantages:

The fermentation mash need not be mixed with a strong mineral acid toliberate the lactic acid, which considerably reduces the salt contentsof the mash. Compared with methods in which strongly acidic cationexchangers are used to separate the lactic acid from the usuallyacidified liquor by classic ion exchange, the chief advantage consistsin that the amount of mineral acid required for regenerating a weaklyacidic ion exchanger is reduced by roughly two thirds.

The advantage compared with IEC methods with basic anion exchangersconsists in that the lactic acid fraction is obtained as a sharpsymmetrical peak without "tailing".

Further, the separation of lactic acid from other present organic acids,in particular ethanoic acid, is also possible on strongly acidic ionexchangers. Accordingly, pharmaceutical-grade lactic acid withsufficient color stability can be obtained so that it can also be usedfor polymerization.

In contrast to IEC methods with basic exchangers requiring diluted acidsfor elution, pure water can be used as eluting agent in the presentprocess.

The fermentation solution from which the lactic acid is to be separatedmust have a pH greater than 5.0 and accordingly still lies within theworking range of strongly acidic cation exchangers.

The resin located in one or more "preliminary columns" is a cationexchanger, preferably a weakly acidic cation exchanger in H⁺ form, sincethis enables regeneration into H⁺ form with practically theoreticalyields of acid (FIG. 7).

The temperature of the "preliminary column" should be at least 50° C.,but preferably 70°-80° C., since this increases the acidity andaccordingly also the salt separation capacity of weakly acidic cationexchangers and substantially improves the useful capacity (break-pointor breakthrough capacity) of the exchanger.

The strongly acidic cation exchanger found in one or more "separationcolumns" is present in H⁺ form so that a chromatographic separation ofthe lactic acid from other organic acids, in particular ethanoic acid,is also possible.

Upon contact with the strongly acidic cation exchanger, the decationizedfermentation solution is divided into a raffinate fraction I (initialfraction) containing the components without a lactic acid content, aproduct fraction containing lactic acid, and a raffinate fraction II(final fraction) containing primarily ethanoic acid.

Pure, preferably deionized, water is used as eluting agent for washingthe individual fractions out of the separation columns.

The elution temperature lies between room temperature and the stabilitythreshold temperature of the resins employed, preferably between 50° C.and 65° C., so as to prevent microbial infection and increase the numberof theoretical plates.

Only the weakly acidic cation exchanger located in the "preliminarycolumns" is regenerated with a diluted strong mineral acid, preferably a1-2N sulfuric acid, after being completely loaded by the cations of thefermentation solution.

The diluted salt solution occurring in the regeneration of the resins inthe preliminary columns is separated into the corresponding acids andbases by salt-hydrolyzing electrodialysis and fed back to the process.

BRIEF DESCRIPTION OF THE DRAWING

The above and other objects, features and advantages of the presentinvention will now be described in more detail with reference to theaccompanying drawing in which:

FIGS. 1 and 4 are graphical illustrations of IE chromatography of "feed"solution 1 with a strongly acidic cation exchanger;

FIGS. 2 and 3 are graphical illustrations of IE chromatography of "feed"solution 2 with a strongly acidic cation exchanger;

FIG. 5 is a graphical illustration of IE chromatography of "feed"solution 2 with a weakly and a strongly acidic cation exchanger;

FIG. 6 is a graphical illustration of IE chromatography of "feed"solution 2 with two strongly acidic cation exchangers;

FIG. 7 is a graphical illustration of regenration of a weakly acidiccation exchanger;

FIG. 8 is a graphical illustration of regenration of a strongly acidiccation exchanger; and

FIG. 9 shows a schematic view of a semicontinuous chromatographyinstallation for extraction of pure lactic acid with "preliminarycolumns" and "separation columns" which are arranged and connectedaccording to the principle of the "simulated moving bed" process.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

More particularly, the process according to the invention is carried outin such a way that a lactic acid mash which is preferably produced byfermentation with lactobacillus delbrueckii with ammonia water asneutralizing agent is freed from coarsely dispersed components(biomass). This is effected, e.g., by centrifuging with a solid-jacketcentrifuge. The fermentation solution may be mixed with activatedcharcoal to remove coloring matter and lipophilic components. After aperiod of contact, the activated charcoal is removed in conventionalmanner by filtration. The mash is concentrated, as the case may be, to30-50% (w/w) by vacuum evaporation and provides the solution which isreferred to hereinafter as "feed".

The actual purification and separation process of the lactic acidaccording to the invention is effected in two steps:

1) by a genuine ion exchange on a preferably weakly acidic cationexchanger in H⁺ form, and

2) by an IE chromatography process on a strongly acidic cation exchangerin H⁺ form.

The process begins by applying a determined volume of fermentationsolution ("feed") to the "preliminary column". The amount of "feed"solution to be applied per chromatography cycle depends on itsconcentration and on the diameter of the "separation columns". Ingeneral, however, the amount lies between 5% and 10% of the resin bedvolume of the "separation column".

When the "feed" solution has fully penetrated into the "preliminarycolumn", pure, preferably deionized, water, the eluting agent used inthis process, is switched to immediately.

A preferably weakly acidic cation exchanger in H⁺ form is located in the"preliminary column". The salts contained in the "feed" solution,primarily the salt of lactic acid (NH₄ lactate), are converted into thecorresponding acids by exchange of ions in this cation exchanger. Theresin bed volume of the "preliminary column" need only be large enoughso that, under the given operating conditions, the so-called"breakthrough capacity" of the resin used in the "preliminary column" issufficient for the amount of cations applied per chromatography cycle tobe exchanged with H⁺ ions. Since the "breakthrough capacity" of weaklyacidic exchangers depends not only on the pH of the "feed" solution butalso, above all, on the operating temperature, it is advantageous toallow the ion exchange process in the "preliminary column" to take placeat a temperature of at least 50° C., but preferably at 70°-80° C.

The decationized "feed" solution coming from the "preliminary column" isthen conveyed via a pump directly to the "separation column" which ispacked with a strongly acidic cation exchanger in H⁺ form. The optimumflow rate required for the chromatography process is also maintainedconstant by means of the pump. The actual IEC process resulting in theseparation of the lactic acid from the rest of the components in the"feed" solution takes place in the "separation column". The eluateflowing out of the "separation column" can be divided into threefractions:

*) the raffinate fraction I (initial fraction) containing the componentsof the "feed" solution other than lactic acid (carbohydrates, strongacids, proteins)

*) the product fraction containing the free lactic acid, and

*) the raffinate fraction II (last fraction) containing primarilyethanoic acid.

The IEC process in the "separation columns" preferably takes place at atemperature of 50°-65° C. This prevents microbial infection of the resinand increases the number of theoretical plates.

The weakly acidic cation exchanger located in the "preliminary column"is regenerated with a strong mineral acid (e.g., with a 1-2N sulfuricacid) by the countercurrent method after being loaded by the cations ofthe "feed" solution. The diluted saline solution which occurs duringregeneration of the "preliminary column" and which is principallycomposed of ammonium sulfate, can be separated by salt-hydrolyzingelectrodialysis into the corresponding acids (sulfuric acid) and bases(ammonium hydroxide) and fed back into the process.

As was already mentioned, the optimum pH value for the microorganismmust be maintained constant during the fermentation of the lactic acidin order to achieve satisfactory productivity. This is effected in asimple manner by adding a neutralizing agent such as NH₄ OH to the mashduring fermentation so that the pH of the solution is held constant at5.5-6.5. Thus, in addition to small amounts of salts, carbohydrates andother impurities stemming from the nutrients, the mash coming out of thefermentation contains the salt of lactic acid, e.g., NH₄ lactate, as themain component.

A number of effects come into play in chromatographic separation ofmulticomponent systems such as the lactic acid mash described above.

The first effect is the "ion exclusion effect" described by WHEATON andBAUMANN (1953) (Ing. Eng. Chem.; 45 (1953) 228). This enables theseparation of ionic and nonionic components. In this case, as opposed tothe conventional ion exchange process, there is no exchange of ions, sothat regeneration of the resin is also not required. The theory of theion exclusion effect can be explained and described by the "Donnanmembrane theory" (MEYER, W. R. S., et al.; Ind. Eng. Chem. Proc. Des.Develop.; 6 (1967) 55). For example, if a salt solution K⁺ A is broughttogether with a cation exchange resin which has been saturatedbeforehand by the cation K⁺, the resin takes on a negative charge inrelation to the solution. This negative charge is the result of a slighttraveling of the anion A in the resin and a similar traveling of thecation K⁺ in the intermediate space of the resin. A potential differencedevelops between the solution and the resin phase. The magnitude of thispotential difference can be described by the Donnan membrane potential.

For practical applications this means that dissolved salts which, ascations, contain the same ion as the counter-ion of the resin cannotpenetrate into the resin and will be the first component to exit thecolumn. The same principle applies also to acids to be separated on astrongly acidic cation exchanger in H⁺ form. However, in the case ofacids, the pKs value of the acid in question still plays a part. Thelower the pKs of the acid (i.e., the stronger the acid), the less theacid will be retained by the resin, and vice versa. This means thatstrong acids cannot penetrate into the resin pores due to Donnanexclusion, whereas medium-strong and weak acids, depending on theirdegree of dissociation, can penetrate to a varying extent into the resinpores and therefore exit the column only later on.

Another effect coming into play in IE chromatography is the so-called"molecular sieve effect" which is responsible for the separation ofuncharged (nonionic) dissolved components (WALTER, H. G. et al., Cer.Sci. Today; 15 (1970) 140). The pore size of an ion exchange resin isdetermined by its degree of cross-linking. The size and shape of amolecule, as much as the pore size of the resin, determines whether amolecule can move from the intermediate space into the pore space whentraveling through an ion exchange column. The ion exchanger acts as amolecular sieve which excludes molecules of a determined shape and size(NORMANN, L. G.; J. Amer. Soc. Sugar Beet Tech.; 12 (1963) 363). Wheneluting the column, those components which cannot move into the poreswill appear in the eluate sooner than those which are uniformlydistributed.

The so-called "distribution coefficient" (K_(di)) provides a yardstickfor determining the degree to which a substance will be held back by theresin in question. It is defined as the ratio of the concentrations ofthe dissolved substance (i) in the resin pore volume (C_(hi)) to theconcentration of the dissolved substance (i) in the intermediate spacevolume (C_(zi)):

    K.sub.di =C.sub.hi /C.sub.zi.

Very generally, the distribution coefficient for a determined compounddepends on the structure of this compound and its concentration, on thetype and ionic form of the resin and, finally, also on the othercompounds present in the solution.

The chromatographic separation of lactic acid from a solution containingnonionic components (carbohydrates) in addition to ionic components(salts) is carried out on a strongly acidic cation exchanger by acombination of the two effects described above. However, a preconditionfor this consists in that there be no reversal of charge in the resinduring the chromatography process, i.e., that the counter-ion of thecation exchanger must be the same as the cation most present in thesolution in terms of quantity. For example, when the mash is neutralizedwith NH₄ OH, this would be the NH₄ ⁺ ion.

In a purely theoretical respect, a chromatographic separation of the NH₄lactate from the rest of the components in the mash would have to bepossible on a strongly acidic cation exchanger in NH₄ ⁺ form. However,experiments have shown that a separation of the lactic acid salt with acation exchanger in this charge form is only possible, if at all, withgreat difficulty, since the distribution coefficients of the individualcomponents are not sufficiently different from one another in this resinform in the sense indicated above (Example 1, FIG. 1).

Moreover, a process such as this would not yield the free acids, butrather, naturally, the salt of the lactic acid, i.e., the NH₄ lactate,which would first have to be converted into the free acid in asubsequent step.

On the other hand, by carrying out the IEC process on a strongly acidiccation exchanger in H⁺ form with a mash whose pH value has beendecreased to 2.5 by a strong mineral acid (e.g., with concentrated H₂SO₄), a totally different chromatogram would result. The acids aredisplaced from their salts by acidifying the mash. The main componentsfound in a solution of this kind are, aside from free lactic acid,primarily (NH₄)₂ SO₄ and sulfuric acid. During chromatographicseparation of this solution, the ammonium sulfate entering theseparation resin along with the "feed" would lead to a partial chargereversal in the resin. The occurring sulfuric acid, along with thesulfuric acid which is present anyway in the "feed", will travel throughthe "separation column" most quickly in accordance with the "ionexclusion effect" and will also be the first component to exit thecolumn. On the other hand, the lactic acid with a much weaker degree ofdissociation (pKs 3.86) will travel more slowly through the separationcolumn and, depending on its strength, will return a portion of thepreviously NH₄ ⁺ -charged resin into its H⁺ form. The lactic acidappears as a double peak in the chromatogram, where the first peak showsthe NH₄ lactate and the second peak shows the free lactic acid (Example2, FIG. 2).

However, the double peak, that is, the division of the lactic acidpresent in the "feed" into a salt fraction and an acid fraction, can beprevented when a determined amount of preferably 2N H₂ SO₄ is fedimmediately or at a determined interval after the charge of "feed"solution which is acidified to a pH of 2.5. The amount of sulfuric acidwhich must be fed in this way is determined by the NH₄ ion concentrationof the "feed" solution. After the sulfuric acid is fed, elution iscarried out, as is conventional, with deionized water. The free lacticacid appears as a sharp single peak in the chromatogram. The reason forthis again is that the strong sulfuric acid travels through theseparation resin much faster than the weaker lactic acid or than themuch weaker ethanoic acid. The sulfuric acid charged after the "feed"solution will, as it were, "outrun" the lactic acid and other weak acidsand will cancel the charge reversal of the resin into its NH₄ ⁺ formcaused by the ammonium sulfate (Example 3, FIG. 3).

A combination of ion exchange and IE chromatography already takes placein this process, albeit in the same column and in the same resin.However, the displacement of the lactic acid from its salt can also beeffected directly by ion exchange on the separation resin. If a mashwhich is not acidified (pH 5.8) is applied as "feed" to a stronglyacidic cation exchanger in H⁺ form and followed in the same way asdescribed above by a determined amount of sulfuric acid and is elutedwith deionized water, the same chromatogram will result (Example 4, FIG.4).

However, the process described above in which both "ion exchange" and"IE chromatography" take place on one and the same resin, a stronglyacidic cation exchanger in H⁺ form, has a decisive disadvantage. Thisconsists in that the amount of sulfuric acid which must be fed after the"feed" solution cannot conceivably be sufficient in practice for acomplete regeneration of the cation exchanger. In each cycle, adetermined amount of NH₄ ions will remain on the exchanger. This meansthat the exchanger must be completely regenerated after a determinednumber of IEC cycles.

This disadvantage may be overcome in a simple manner by allowing the twoprocesses, "ion exchange" and "IE chromatography", to run separately.For this purpose it is necessary that a "preliminary column" in whichthe cations in the "feed" solution are exchanged for H⁺ ions by ionexchange is arranged upstream of the "separation column" which is packedwith the strongly acidic cation exchanger in H⁺ form and in which onlythe IEC process will take place. Therefore, this "preliminary column" islikewise packed with a cation exchanger in H⁺ form which can have astrongly acidic as well as a weakly acidic character. However, since thepH value of the fermentation mash is greater than 5 and accordinglystill lies in the working range of weakly acidic cation exchangers andhas an acidity sufficient at least to hydrolyze the salt of a weak acid(lactic acid), it is advantageous in the process according to theinvention to use weakly acidic cation exchangers in the "preliminarycolumn". The resin bed volume of the "preliminary column" depends on the"feed" quantity applied per chromatography cycle and on the"breakthrough capacity" of the resin. Although weakly acidic cationexchangers have a very high total capacity (up to 4 val/l resin), theiruseful capacity (breakthrough capacity) is reduced by operatingconditions. Aside from the pH value of the solution (insofar as it liesbelow 7 or lies between 7 and 4), the operating temperature has thegreatest influence on the useful capacity of weakly acidic cationexchangers. Thus, the acidity and accordingly the salt-hydrolyzingcapacity of the carboxylic acid resin, also relative to neutral salts,at first increases only slightly with increasing temperature, butincreases very sharply above 50° C. Therefore, it is necessary tooperate the "preliminary column" at the highest possible temperature,preferably 70° C.-80° C. (Examples 5 and 6, FIGS. 5 and 6).

As will be seen from FIGS. 5 and 6, an extensive separation of lacticacid can be achieved already with the separating distance of roughly 800cm available in the experimental facility. However, following the law of"theoretical plates", a complete separation of the lactic acid is to beexpected with the separating distances of 15 to 20 m conventionally usedin practice. Above all, a complete separation of carbohydrates from thelactic acid can be effected as is demonstrated in the "heat" test shownin Example 5, in which there was no discoloration after heating thelactic acid-containing fraction at 200° C. for one hour.

A decisive advantage of weakly acidic cation exchangers compared withstrongly acidic cation exchangers consists in the high chemicalutilization factor in regeneration. Due to the low dissociationconstant, their regeneration into H⁺ form can be carried out practicallywith the theoretical yield of acid (Examples 7 and 8, FIGS. 7 and 8).

The process described above can be realized in batch operation as wellas in a quasi-continuous apparatus such as that described in the U.S.patent (1961). In this method, in the chromatography process describedin the U.S. patent, the location at which the "feed" stream enters theseparation resin and the location at which the "product" stream exitsfrom the separation resin are constantly changed. This switching processsimulates a double motion of the resin, for which reason it is referredto in the literature as the "simulated moving bed" process. A possiblerealization of this process incorporating the "preliminary columns"required for the process according to the invention is shown in FIG. 9,which shows a schematic view of a semicontinuous chromatographyinstallation for extraction of pure lactic acid with "preliminarycolumns" and "separation columns" which are arranged and connectedaccording to the principle of the "simulated moving bed" process.

In principle, the separation process and purification process arecarried out in this installation precisely as described in thepreceding, with the exception that semicontinuous operation can becarried out in this case. Here, also, a determined "feed" amount isapplied to one of the "preliminary columns" and is rinsed down withwater. After a certain period, an additional "feed" amount is applied tothe next "preliminary column", thus producing a continuous "feed"stream. The decationized "feed" coming from the "preliminary column" isconveyed directly to a determined "separation column" by a pump (P),while the resin in this "preliminary column" is regenerated withsulfuric acid. This cyclical loading and regeneration of the"preliminary columns" can be repeated as often as desired.

The "separation columns", on the other hand, are connected via apipeline to form a ring. This ring line is connected between the columnsvia valves to four supply lines. These supply lines enable four processflows:

*) the "feed" inlet flow

*) the "eluent" inlet flow

*) the "product" outlet flow

*) the "raffinate" outlet flow.

The flow in the installation is maintained constant by an individualpump. In this process, there are always exactly four valves opened,these valves dividing the ring of columns into four zones:

The first zone is the so-called "adsorption zone" and lies between thepoint where the "feed" flow enters the "separation columns" and thepoint where the faster flowing substances are decanted as the"raffinate" flow.

The second zone is the so-called "purification zone" and lies betweenthe point at which the "raffinate" flow exits the "separation columns"and the point where pure water enters the "separation columns" as"eluent" flow.

The third zone is the so-called "desorption zone" and lies between thepoint at which the "eluent" flow enters the "separation columns" and thepoint where the lactic acid is decanted as the "product" flow.

The ethanoic acid and other slow-running components (raffinate fractionII) are likewise decanted with the "product" flow, but are collectedseparately.

The fourth zone is the so-called "buffer zone" and lies between thepoint at which the "product" flow exits the "separation column" and thepoint where the "feed" flow re-enters the separation system.

At given times, these zones are switched one column farther in the flowdirection by means of a process control computer. These switchingprocesses simulate a resin flow in the opposite direction with respectto flow.

Compared to batch operation, the "simulated moving bed" process has theadvantage that roughly one third less resin and roughly two thirds lesseluent is needed for production of the same quantity of lactic acid.

The invention will be explained more fully in the following by examples.The resins used are common commercial products. Their generalcharacteristics as specified by the manufacturer are compiled in Table1:

                                      TABLE 1                                     __________________________________________________________________________                                 resin in separation                                                           columns                                                    resin in preliminary columns                                                                     Dowex Mono                                       feature   Dowex MWC-1                                                                            Lewatit MDS 1368                                                                        C 356 CA                                         __________________________________________________________________________    manufacturer                                                                            Dow Chemical                                                                           Bayer     Dow Chemical                                     resin type                                                                              weakly acidic                                                                          strongly acidic                                                                         strongly acidic                                  polymer base                                                                            polyacrylic                                                                            Styrene/DVB                                                                             Styrene/DVB                                                macroporous                                                                            gel       gel                                              functional group                                                                        carboxyl group                                                                         sulfonic acid                                                                           sulfonic acid                                    particle size                                                                           0.4-1.2 mm                                                                             0.35 mm ± 0.05                                                                       0.35 mm ± 0.05                                moisture content                                                                        44-50 % by wt.                                                                         approx. 49 % by wt.                                                                     57-61 % by wt.                                   bulk density                                                                            720 g/l  830 g/l   833 g/l                                          total capacity                                                                          3.8 val/l                                                                              1.8 val/l 1.5 val/l                                        maximum operating                                                             temperature                                                                             100° C.                                                                         100° C.                                                                          90° C.                                    pH working range                                                                        5-14     1-14      1-14                                             __________________________________________________________________________

The "feed" solution used in all IEC tests was a lactic acid mashproduced by fermentation with lactobacillus delbrueckii. NH₄ OH was usedas a neutralizing agent so that the lactic acid was present in the mashas ammonium lactate. After separation of the biomass by means of achamber centrifuge, the mash was concentrated to approximately 30%wt./vol. by vacuum distillation and then decolorized by adding activatedcharcoal. The pH value in a portion of the mash produced in this way wasreduced to 2.5 by the addition of concentrated sulfuric acid and thelactic acid was accordingly displaced from its salt.

The light-yellow solutions obtained in this way are designated in thefollowing examples as "feed" solution 1 (pH 5.8) and "feed" solution 2(pH 2.5).

Table 2 shows the composition of these solutions determined by highpressure liquid chromatography or HPLC (Shimadzu Co.):

                  TABLE 2                                                         ______________________________________                                        analytical characterization of the fermentation solution by HPLC                           "feed" solution 1                                                                           "feed" solution 2                                               fermentation solu-                                                                          fermentation solu-                                 substance    tion at pH 5.8                                                                              tion at pH 2.5                                     ______________________________________                                        sulfate      5.12 g/l      124.20 g/l                                         oxalic acid  1.07 g/l      0.97 g/l                                           maltose      4.68 g/l      4.25 g/l                                           glucose      1.99 g/l      1.79 g/l                                           mannite      4.64 g/l      4.22 g/l                                           unknown substance A                                                                        2.91 g/l      2.62 g/l                                           unknown substance B                                                                        2.28 g/l      2.05 g/l                                           tactic acid  303.93 g/l    268.40 g/l                                         ethanoic acid                                                                              2.80 g/l      2.52 g/l                                           ______________________________________                                    

EXAMPLE 1

A chromatography testing installation having two double-wall columns ofacrylic glass in series was packed with the strongly acidic cationexchanger DOWEX Mono C 356 CA. The diameter of the inner column was 2.0cm and the length of each column was 200 cm. At a resin bed height of365 cm, this resulted in a resin bed volume of 1150 cm³.

After the columns were packed with the resin, they were first brought toH⁺ form with 2N HCl and then charged to NH₄ ⁺ form with ammonia water(approx. 5%). All of the liquid volumes fed to the columns during theexperiment were degassed beforehand and preheated to the columntemperature. This temperature was 55° C. and was maintained constant bycirculating thermostats.

After feeding 100 ml of the "feed" solution 1 to the first column,elution was carried out with degassed, deionized water which waspreheated to approximately 60° C. The flow rate of 6 ml/min (1.91ml/min*cm²) was adjusted by a constricted-tube or peristaltic pump andmaintained constant. The eluate exiting from the bottom of the firstcolumn was immediately pumped to the second column. The eluate of thesecond column was guided through a conductivity measurement cell andthen collected in a fraction collector. The signal coming from theconductivity measurement cell was recorded by a plotter in the form of aconductivity chromatogram. This served only for rough detection, sinceonly the position of the salts and acids can be detected byconductivity.

A more precise analysis of the collected fractions was carried out byHPLC. In this way the carbohydrates as well as the acids and salts couldbe determined quantitatively.

The concentrations of the individual substances determined by HPLC wereplotted against the eluate volume and resulted in the chromatogram shownin FIG. 1.

EXAMPLE 2

The procedure was identical to Example 1, except that the stronglyacidic cation exchanger was converted to H⁺ form with 2N sulfuric acidand 100 ml of the "feed" solution 2 was applied to the resin. Thechromatogram of this separation is shown in FIG. 2.

EXAMPLE 3

The procedure was identical to Example 2, except that the chromatographyinstallation was supplemented by two additional columns so that a resinbed volume of approximately 2300 cm³ resulted in a resin bed height of730 cm. Only 60 ml of"feed" solution 2 was applied to the resin in thistest. Elution was carried out such that 180 ml of 2N H₂ SO₄ weresupplied after rinsing the "feed" solution with 40 ml water in the resinbed. Next, water was switched to again and elution was brought tocompletion as described in Example 1. The chromatogram of thisseparation is shown in FIG. 3.

EXAMPLE 4

The procedure was identical to Example 3, except that 60 ml of "feed"solution 1 was applied to the resin. The chromatogram of this separationis shown in FIG. 4.

EXAMPLE 5

The procedure was identical to Example 4, except that the chromatographyinstallation was supplemented by a "preliminary column". At a diameterof 2.5 cm and a resin bed height of 80 cm, this "preliminary column" hada resin bed volume of approximately 400 cm³. The "preliminary column"was packed with the weakly acid cation exchanger Dowex MWC-1 and broughtto H⁺ form by a 1N H₂ SO₄. The "preliminary column" was maintained at aconstant temperature of 75° C. by a circulating thermostat. Next, 60 mlof "feed" solution 1 were applied to the "preliminary column" andelution was carried out with preheated (70° C.) deionized water with aflow rate of 9 ml/min (2.8 ml/min*cm²). The chromatogram of thisseparation is shown in FIG. 5.

The lactic acid fraction was subjected to the so-called "heat" test. Forthis purpose, a sample of this fraction was heated for one hour at 200°C. in a glycerin bath in a pressure-proof sealable tube. Extinctionbefore and after heating was determined at 420 nm (layer thickness 1cm).

                  TABLE 3                                                         ______________________________________                                        feature           extinction                                                  ______________________________________                                        "feed" solution 1 0.412                                                       sample before heating                                                                           0.006                                                       sample after heating                                                                            0.014                                                       ______________________________________                                    

EXAMPLE 6

The procedure was identical to Example 5, except that the "preliminarycolumn" was packed with the strongly acidic cation exchanger Lewatit MDS1368 and brought to H⁺ form with H₂ SO₄ (2N) The chromatogram of thisseparation is shown in FIG. 6.

EXAMPLE 7

The weakly acidic cation exchanger Dowex MWC-1 in the "preliminarycolumn" which was loaded with NH₄ ⁺ ions in Example 5 was regeneratedwith H₂ SO₄ (1N). The solution exiting from the "preliminary column" ranthrough a conductivity measurement cell and was then collected in afraction collector. The concentration of ammonium sulfate in theindividual fractions was determined by adding concentrated NaOH and bysubsequent steam distillation with the Buchi "distiller unit". Theresults of this test are shown in FIG. 7.

EXAMPLE 8

The procedure was identical to Example 7, except that the stronglyacidic cation exchanger Lewatit MDS 1368 which was loaded with NH₄ ⁺ions in Example 6 was regenerated in the "preliminary column" with H₂SO₄ (1N) The results of this test are shown in FIG. 8.

We claim:
 1. Process for the separation and purification of lactic acidfrom salt-containing and carbohydrate-containing substrates from afermentation solution from which coarsely dispersed and lipophilicimpurities have been removed, said separation and purification processcomprising the steps of:a) converting the salts which may be present inthe fermentation solution, principally the salt of lactic acid, intofree acids by means of genuine ion exchange in one or more "preliminarycolumns", and b) separating the free lactic acid from the rest of theacids, carbohydrates and other impurities present in the fermentationsolution by chromatography at strongly acidic ion exchangers in one ormore "separation columns".
 2. Process according to claim 1, wherein thefermentation solution from which the lactic acid is separated has a pHgreater than 5.0.
 3. Process according to claim 1 wherein the resinlocated in one or more "preliminary columns" is a cation exchanger. 4.Process according to claim 3 wherein the temperature of the "preliminarycolumn" is at least 50° C.
 5. Process according to claim 4 wherein thetemperature of the "preliminary column" is 70°-80° C.
 6. Processaccording to claim 3 wherein the cation exchanger is a weakly acidiccation exchanger in H⁺ form.
 7. Process according to claim 1, whereinthe strongly acidic cation exchanger found in one or more "separationcolumns" is present in H⁺ form.
 8. Process according to claim 1, whereinthe decationized fermentation solution, upon contact with the stronglyacidic cation exchanger, is divided into a raffinate fraction I (initialfraction) containing the components without a lactic acid content, aproduct fraction containing lactic acid, and a raffinate fraction II(final fraction) containing primarily ethanoic acid.
 9. Processaccording to claim 8 wherein the elution temperature lies between roomtemperature and the stability threshold temperature of the resinsemployed.
 10. Process according to claim 9, wherein the diluted saltsolution occurring in the regeneration of the resins in the preliminarycolumns is separated into the corresponding acids and bases bysalt-hydrolyzing electrodialysis and fed back to the process. 11.Process according to claim 9 wherein the elution temperature liesbetween 50° C. and 65° C.
 12. Process according to claim 1 wherein purewater is used as eluting agent for washing the individual fractions outof the separation columns.
 13. Process according to claim 12 wherein thepure water is deionized.
 14. Process according to claim 1 wherein onlythe weakly acidic cation exchanger located in the "preliminary columns"is regenerated with a diluted strong mineral acid after being completelyloaded by the cations of the fermentation solution.
 15. Processaccording to claim 14 wherein the diluted strong mineral acid is a 1-2Nsulfuric acid.